The overall objective of this proposal, which is a response to the Program Announcement New Directions in Pain Research, is to study the role of peripheral glutamate receptors in the generation of ectopic discharges from injured nerves as it relates to neuropathic pain. Preliminary evidence is presented that glutamate is accumulated at the site of nerve injury and block of ionotropic glutamate receptors decreases ectopic discharges recorded from injured nerve axons and attenuates pain behaviors in a rat model of neuropathic pain. The following hypotheses will be tested: 1. Blockade of ionotropic glutamate receptors or inhibition of glutamate synthesis at the site of nerve injury attenuates neuropathic pain states; 2. Peripheral nerve injury elicits an augmented expression of ionotropic glutamate receptors at the site of nerve injury and an increased glutaminase activity in the neuroma, dorsal root ganglia and spinal cord; 3. Peripheral nerve injury increases local glutamate accumulation, which is attenuated by inhibition of glutaminase or blockade of ionotropic glutamate receptors; and 4. Glutamate accumulation at the site of nerve injury contributes to the generation of abnormal afferent discharges, which play an important role in the sensitization of spinal dorsal horn neurons. A tight partial sciatic nerve ligation rat model of neuropathic pain will be used. Behavioral assessment of allodynia will be threshold to withdrawal using von Frey filaments. Motor function will also be evaluated. Ectopic nerve activity proximal to the ligation will be measured by extracellular recording from split filaments and assigned to fiber class on the basis of conduction velocity. Single unit recording of dorsal horn neural activity will also be done. Immunocytochemistry and electron microscopy will be used to detect changes in glutamate receptor expression, which will also be measured quantitatively by Western blot. Glutamate and aspartate content of nerve samples will be measured with HPLC. Glutaminase will be measured by HPLC determinations of glutamate over time.
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